Kolarov J, Kolarova N, Nelson N
Roche Institute of Molecular Biology, Roche Research Center, Nutley, New Jersey 07110.
J Biol Chem. 1990 Jul 25;265(21):12711-6.
The op1 mutation in yeast is known to be due to a defect in the mitochondrial ADP/ATP translocator. Sequencing of the gene AAC2 revealed that the mutation resulted from a single base change that caused a replacement of arginine 97 by a histidine. The gene encoding AAC2 was also cloned and sequenced from an op1 revertant capable of growth on glycerol as a sole carbon source. Sequence analysis indicates that the reverted gene underwent rearrangement in which a portion of an unknown gene was used to repair the mutation. An oligonucleotide complementary to this insert was used to clone a previously unrecognized gene encoding ADP/ATP translocator in yeast. The newly discovered gene, AAC3, is homologous with the previously known genes AAC1 and AAC2. Gene disruption experiments suggest that AAC2 encodes the majority of the translocator. Expression of AAC1 and AAC2 required derepressed conditions whereas expression of AAC3 occurred almost exclusively under anaerobic conditions. Both the op1 mutant and the strain that contains an interrupted AAC2 were able to grow under anaerobic conditions, suggesting that AAC3 can replace the gene product of AAC2. Indeed, when cloned into multicopy plasmid, AAC3 was able to replace the disrupted AAC2 in the JLY-73 strain. The concomitant disruption of the AAC2 and AAC3, however, results in arrest of cell growth under conditions of low oxygen tension. The discovery of a third gene encoding ADP/ATP translocator helps to clarify certain characteristics of op1 mutants which could not be resolved in the past.
已知酵母中的op1突变是由于线粒体ADP/ATP转位酶存在缺陷所致。对AAC2基因进行测序后发现，该突变是由单个碱基变化引起的，导致第97位精氨酸被组氨酸取代。还从能够以甘油作为唯一碳源生长的op1回复突变体中克隆并测序了编码AAC2的基因。序列分析表明，回复后的基因发生了重排，其中一个未知基因的一部分被用于修复突变。与该插入片段互补的寡核苷酸被用于克隆酵母中一个先前未被识别的编码ADP/ATP转位酶的基因。新发现的基因AAC3与先前已知的基因AAC1和AAC2同源。基因破坏实验表明，AAC2编码了大部分的转位酶。AAC1和AAC¬2的表达需要去阻遏条件，而AAC3的表达几乎只在厌氧条件下发生。op1突变体和含有中断的AAC2的菌株在厌氧条件下都能够生长，这表明AAC3可以替代AAC2的基因产物。事实上，当克隆到多拷贝质粒中时，AAC3能够替代JLY - 73菌株中被破坏的AAC2。然而，AAC2和AAC3的同时破坏会导致在低氧张力条件下细胞生长停滞。第三个编码ADP/ATP转位酶的基因的发现有助于阐明op1突变体的某些过去无法解决的特征。
